Study On The Quality Attributes Of Shengjiang Powder And Its Effect And Mechanism On Improving Acute Lung Injury In Mice

Objective:Shengjiang Powder is derived from the Typhoid Plague Identification of Yang Lishan,a doctor in the Qing Dynasty.Based on the theory of traditional Chinese medicine,this paper established the preparation process of Shengjiang Powder,conducted experimental research on the quality attributes of Shengjiang Powder,and established the content determination method and fingerprint spectrum method of Shengjiang Powder,so as to formulate a comprehensive and overall quality control and evaluation method for Shengjiang Powder.Then,based on network pharmacology,we established acute lung injury model to verify the rationality and quality effectiveness of Shengjiang Powder.At the same time to explore the pharmacodynamic effect of Shengjiang Powder in the therapy of acute lung injury（ALI）,and to provide theoretical foundations for the developing and research of Shengjiang Powder.Methods:（1）By consulting medical classics and literature,the origin of prescription,the dose of prescription,usage and dosage,efficacy and indications,and the origin of medicinal materials were verified.According the records of ancient books and combining with modern actual production,the amount of wine added in the processing of bombyx batryticatus was preliminarily determined by single factor test,and the processing technology of bombyx batryticatus was further optimized by L₉（3⁴）orthogonal test.The moist heat sterilization method was selected to inspect the pretreatment sterilization of medicinal materials.The preparation process of Shengjiang Powder was determined by investigating the prescription particle size and powder yield,and the rationality of the production process was verified by pilot production.（2）The quality attributes of Shengjiang Powder were explored by microscopic identification,thin layer identification and high performance liquid chromatography,and the fingerprint spectrum of Shengjiang Powder was established to formulate a comprehensive and effective quality standard.（3）TCMSP and chemical database of Chinese Academy of Sciences were used to search effective components of Shengjiang Powder.Gene Cards database was used to acquire the disease targets of ALI.Cytoscape 3.7.2 software was used to construct the relevant between component and disease network diagram.STRING database was used to serach the main targets to construct PPI network.DAVID website was used to analyze the GO and KEGG enrichment of key target proteins.A mouse model of LPS-induced acute lung injury was established to explore the pharmacodynamic effects of Shengjiang Powder and decoction.The lung wet weight index and lung pathology were compared among different groups.We tested the serous changes of interleukin-1β,interleukin-6 and tumor necrosis factor-αby the way of enzyme-linked immunosorbent assay.The expressions of TLR4,p38-MAPK,ERK and JNK-1 mRNA were checked by qRT-PCR.Western Blot was used to check out the expressions of nuclear factor-kappa B,phosphorylated NF-κB and nuclear factor-κB inhibitorαin lung.Results:（1）Shengjiang Powder is an oral powder evolved from Neifuxianfang and Peizhensan.The medicinal materials are confirmed to be consistent with the varieties included in the Chinese Pharmacopoeia after textual research.Determine the processing parameters of stiff silkworm with wine:100 gram of stiff silkworm with wine was mixed with 12 gram of yellow rice wine,moistened for 45 minutes,fried at 120 degree celsius for25 minutes,fried yellow,removed and dried.The preparation process of Shengjiang Powder was determined as follows:274 gram of bombyx batryticatus,137 gram of cicadae periostracum,41 gram of curcumae longae rhiaoma and 548 gram of rhei radix et rhizome were taken.The above four herbs were sterilized by damp-heat sterilization（121 degree celsius,30 minutes）,crushed into fine powder after drying,sieved and mixed.（2）The microscopic identification method was used to identify the characteristics of bombyx batryticatus,cicadae periostracum,curcumae longae rhiaoma and rhei radix et rhizome in Shengjiang Powder;the specificity identification of curcumae longae rhiaoma and rhei radix et rhizome in Shengjiang Powder was established by thin layer chromatography.The content determination method of total anthraquinones in rhei radix et rhizome in Shengjiang Powder was established by high performance liquid chromatography.The results of methodology and system adaptability were in line with the requirements of pharmacopoeia,indicating that this method can be used for the determination of total anthraquinones in rhubarb in Shengjiang Powder.The HPLC fingerprint of 10 batches of Shengjiang Powder was established,and there were 9 common peaks were marked.Through the investigation of precision,repeatability and stability,it was determined that the determination method met the requirements of characteristic chromatogram,and the method was stable and feasible.（3）The component-target-disease network was constructed by network pharmacology,and we have predicted 95 potential targets for the treatment of acute lung injury.The key targets such as TNF,TP53,MAPK3,IL-1βand EGFR were found by protein interaction network.The results of enrichment analysis showed that the key targets of Shengjiang Powder in the treatment of acute lung injury（ALI）mainly exerted anti-inflammatory effects through multi-pathway synergy,such as PI3K-Akt signaling pathway,p53 signaling pathway,NF-κB signaling pathway,MAPK pathway,etc.The pharmacodynamics of Shengjiang Powder in the treatment of ALI was explored through in vitro experiments.After modeling,the lung index of mice increased significantly,a large number of inflammatory cells infiltrated in the alveolar cavity,alveolar septum and alveolar wall thickened significantly,and some alveolar cavities collapsed.The expression levels of TNF-α,IL1βand IL-6 were tested by ELISA.Compared with the model group,the levels of inflammatory cytokines in the treatment group were significantly lower（P<0.05）.The results of qRT-PCR showed that compared with the model group,the expression levels of TLR4 mRNA,p38-MAPK,ERK and JNK-1mRNA in each administration group showed a downward trend;Western Blot results showed that compared with the model group,the administration of each drug.Conclusion:Through the Chinese medical classics and literature resources,prescription composition and prescription dose were determined.The processing method of wine bombyx batryticatus was established,and the preparation process of Shengjiang Powder traditional powder was determined.At the same time,the research method of the quality attributes of Shengjiang powder was established.The total quality control and evaluation were carried out by microscopic identification,thin layer identification,total anthraquinone content determination and characteristic spectrum method.The pharmacological model of acute lung injury in mice was established by network pharmacology,and the efficacy of powder and decoction was compared.It is preliminarily verified that ALI can play an anti-inflammatory role through TLR4/NF-κB,NF-κB/IL-6,MAPK and other signaling pathways,and the treatment effect of high dose powder is better than that of decoction.